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Title: Structural basis for high substrate-binding affinity and enantioselectivity of 3-quinuclidinone reductase AtQR

Highlights: • Crystal structure of AtQR has been determined at 1.72 Å. • NADH binding induces the formation of substrate binding site. • AtQR possesses a conserved hydrophobic wall for stereospecific binding of substrate. • Additional Glu197 residue is critical to the high binding affinity. - Abstract: (R)-3-Quinuclidinol, a useful compound for the synthesis of various pharmaceuticals, can be enantioselectively produced from 3-quinuclidinone by 3-quinuclidinone reductase. Recently, a novel NADH-dependent 3-quinuclidionone reductase (AtQR) was isolated from Agrobacterium tumefaciens, and showed much higher substrate-binding affinity (>100 fold) than the reported 3-quinuclidionone reductase (RrQR) from Rhodotorula rubra. Here, we report the crystal structure of AtQR at 1.72 Å. Three NADH-bound protomers and one NADH-free protomer form a tetrameric structure in an asymmetric unit of crystals. NADH not only acts as a proton donor, but also contributes to the stability of the α7 helix. This helix is a unique and functionally significant part of AtQR and is related to form a deep catalytic cavity. AtQR has all three catalytic residues of the short-chain dehydrogenases/reductases family and the hydrophobic wall for the enantioselective reduction of 3-quinuclidinone as well as RrQR. An additional residue on the α7 helix, Glu197, exists near the active site ofmore » AtQR. This acidic residue is considered to form a direct interaction with the amine part of 3-quinuclidinone, which contributes to substrate orientation and enhancement of substrate-binding affinity. Mutational analyses also support that Glu197 is an indispensable residue for the activity.« less
Authors:
;  [1] ;  [2] ;  [3] ;  [1] ;  [4] ;  [5] ;  [2] ;  [3] ;  [1] ;  [4] ;  [3] ;  [1]
  1. Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657 (Japan)
  2. Division of Applied Life Sciences, Graduate School of Life and Environmental Sciences, Osaka Prefecture University, 1-1 Gakuen-cho, Naka-ku, Sakai 559-8531 (Japan)
  3. (Japan)
  4. Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-Oiwakecho, Sakyo-ku, Kyoto 606-8502 (Japan)
  5. Research and Development Center, Nagase and Co., Ltd., 2-2-3 Muratani, Nishi-ku, Kobe 651-2241 (Japan)
Publication Date:
OSTI Identifier:
22416379
Resource Type:
Journal Article
Resource Relation:
Journal Name: Biochemical and Biophysical Research Communications; Journal Volume: 446; Journal Issue: 4; Other Information: Copyright (c) 2014 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved.; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; AFFINITY; AMINES; CRYSTAL STRUCTURE; DRUGS; NADH2; OXIDOREDUCTASES; PROTONS; REDUCTION; RESIDUES; STABILITY; SUBSTRATES; SYNTHESIS